Each year, seasonal influenza causes over 300,000 hospitalizations and 36,000 deaths in the US alone (Simonsen et al., Lancet Infect Dis 7:658-66, 2007). The emergence of the novel H1N1 influenza virus in 2009 demonstrated how quickly a new influenza pandemic can sweep across the world. The spread of highly pathogenic H5N1 viruses in birds and coincident infections in humans have raised the concerns that H5N1 viruses may cause a new pandemic in humans. Vaccination is an effective method to prevent influenza infection. There are two influenza vaccine approaches licensed in the United States; the inactivated, split vaccine and the live-attenuated virus vaccine. Inactivated vaccines can efficiently induce humoral immune responses but generally only poor cellular immune responses.
Baculoviruses are a family of large rod-shaped enveloped viruses with a large circular double-stranded DNA genome (80-200 kb). Baculoviruses infect some insects, but not mammals (Blissard, Cytotechnology 20:73-93, 1996). Autographa californica multiple nucleopolyhedrovirus (AcMNPV) is the most well studied baculovirus and most extensively used for protein expression because the polyhedrin (PH) and p10 promoters are efficient promoters (McMichael et al., N Engl J Med 309:13-17, 1983). Compared to other protein expression approaches, the baculovirus expression vector system (BEVS) produces abundant protein yields with appropriate eukaryotic glycosylation and other modifications. This system has also been used for virus-like particle (VLP) production for vaccines against HIV, HPV and influenza (Gheysen et al., Cell 59:103-112, 1989; Kirnbauer et al., J Virol 67:6929-6936, 1993; Latham et al., J Virol 75:6154-6165, 2001). However, the baculovirus (BV)-derived VLPs are always accompanied with BV contamination. Therefore, separating VLPs from contaminating BV is an obstacle that needs to be overcome.
Due to its low cytotoxicity and absence of pre-existing antibodies (Kost et al., Nat Biotechnol 23:567-575, 2005; Strauss et al., Mol Ther 15:193-202, 2007), AcMNPV has emerged as a potent vaccine vector (Fan et al., J Virol Methods 150:21-26, 2008; Feng et al., DNA Cell Biol 25:668-673, 2006; Lin et al., Vaccine 26:6361-6367, 2008; Prabakaran et al., Virology 380:412-420, 2008; Yoshida et al., Infect Immun 77:1782-1789, 2009). Foreign immunogens or peptides can be displayed on the envelope of AcMNPV by fusion with the baculovirus major envelope protein gp64 (Boublik et al., Biotechnology (NY) 13:1079-1084, 1995; Oker-Blom et al., Brief Funct Genomic Proteomic 2:244-253, 2003).